The use of alpha amino acids has recently undergone substantial development because of new uses uncovered in the areas of medicine and food. The preparation of L-alpha amino acids has become increasingly important particularly in view of the fact that the L-alpha amino acids have been shown in some instances to be more effective than the D-alpha amino acids. Further, with the development of the artificial sweetener aspartame, an increasing need has arisen for a precursor, L-phenylalanine.
Since racemates of D,L-alpha amino acids contain one-half of either isomer, resolution of the racemates into one isomer can have a theoretical yield of only 50 percent. The profitability of any resolution method is directly tied to methods for racemizing the residue left after resolution into a D,L-racemate for further resolution. Since resolution is generally the result of selectively changing a derivative of the desired isomer of the amino acid to the desired amino acid isomer, racemization of the remaining isomer derivative must be under conditions that allow formation of a racemate of the derivative for further resolution.
Many methods are known for racemization of free amino acids or their salts such as the aqueous system shown in U.S. Pat. No. 4,401,820. The patent teaches the use of an aldehyde and a glacial acetic acid-water mixture to effect racemization of amino acids. The patent does not provide direction for racemization of amino acids wherein the carbonyl group is substituted. The use of the aqueous media disclosed in this reference can also result in the hydrolysis of some amino acid derivatives, notably esters, resulting in the formation of an amino acid contaminated with aldehyde and acetic acid.
Very few methods are available for racemization of carbonyl-substituted amino acids which allow for the racemization without destruction of the carbonyl-substituent group. Of particular importance is European Patent Publication No. 0089,886 which teaches the use of an insolubilized pyridoxal-5-phosphate to racemize amino acid esters in aqueous media. The essential teaching of the reference is the ability to carry out the chemical catalytic racemization under the same reaction conditions of solvent, pH and temperature as that used for the enzymatic hydrolysis resolution. This patent teaches that an immobilized pyridoxal-5-phosphate can be used to racemize D-phenylalanine ester to the corresponding D,L-racemate in an aqueous solution at pH 7 and a temperature of preferably 20.degree. to 40.degree. C. This is in line with the fact that pyridoxal-5-phosphate is a biological component whose reactions are normally conducted in aqueous medium.
It is known that racemates of D,L-alpha amino acid ester can be hydrolytically resolved into the L-amino acid by use of various esterase enzymes such as chymotrypsin. This enzyme specifically forms the basis of a process for the enzymatic resolution of racemic mixtures of phenylalanine (U.S. Pat. No. 3,813,317). In this patent, chymotrypsin is used to hydrolytically resolve an aqueous racemic mixture of ring-substituted phenylalanine esters. The reaction is carried out in an aqueous solution.
The combination of an aliphatic acid and an aldehyde or ketone as a racemizing agent for esters of amino acids is disclosed in co-pending U.S. patent application Ser. No. 642,212 filed Aug. 17, 1984, now abandoned.
In co-pending U.S. patent application Ser. No. 641,889 filed Aug. 17, 1984, now abandoned, there is disclosed a process for racemizing derivatives of amino acids by heating the amino acid derivatives with pyridoxal-5-phosphate in an organic solvent system.
The advantages of using an organic solvent system in combination with pyridoxal-5-phosphate are discussed in the article by Pugniere et al. appearing in Biotechnology Letters 7, 31-6 (1985).
While the use of pyridoxal-5-phosphate in an organic solvent system as a racemizing agent does possess a number of advantages, this method can be expensive due to the cost of the pyridoxal-5-phosphate as well as the fact that upon prolonged storage, the pyridoxal-5-phosphate can exhibit a serious loss of racemizing activity.
Similarly, the racemization of amino acids or derivatives of amino acids in the presence of aliphatic or mineral acids may also present problems. The presence of the mineral acid can lead to the formation of the acid salt of the amino acid which must be first neutralized to destroy all acidity. This adds an extra reaction step and can result in some loss of amino acid. In the presence of aliphatic acids, amino acid esters tend to undergo peptidization unless they are present as the mineral acid salt. When either mineral and/or aliphatic acids are present, an extra neutralization step will be required and waste disposal of the salt formed in neutralization will also be required before any resolution process can be carried out.
Another method for the racemization of amino acids or amino acid derivatives involves the use of aromatic aldehydes combined with polymeric resins. A number of references disclose this method.
Russian Pat. No. 929,629 discloses the preparation of an aromatic aldehyde-containing polymeric resin wherein the aromatic aldehyde-containing polymeric resin is prepared by reacting a chloromethylated styrene-divinylbenzene copolymer with nitrobenzene solvent, salicylaldehyde and zinc chloride.
Previously discussed EPO patent application No. 0089886 discloses the use of pyridoxal-5-phosphate supported on diethylaminoethyl cellulose for the racemization of L-tyrosine methyl ester at pH 7.
In two Russian patents (USSR No. 593,733 and USSR No. 686,754), there is disclosed a racemization process for amino acids which uses an aromatic resin with hydroxy and aldehyde groups. The racemization is done in the presence of cupric ion. Similar to the above procedure are the methods reported in CA 92, 181930w [corresponding to the article appearing in Vysokomol. Sozed. A22(1), 71-6 (1980)], in Bull. Chem. Soc. Jpn. 35, 1422 (1962), in Bull. Chem. Soc. Jpn. 36, 734 (1963), and in Bull. Chem. Soc. Jpn. 36, 739 (1963).
The above methods suffer from the disadvantage of requiring the presence of a metallic ion. Thus, a second step is needed to remove the metallic ion, complete removal of which is not often possible.
Thus, there is a need for a racemization process which can effectively racemize amino acids or amino acid derivatives without the presence of aliphatic acids, mineral acids or metallic ions. Such a process would eliminate extra neutralization steps, negate the possibility of unwanted peptidization reactions and also eliminate the extra step of removal of metallic ions. There is also a need for the promotion of the racemization process so that complete racemization can be carried out in shorter periods of time using recoverable racemization agents which are storage stable with little to no loss in activity for recyclic use.